Two classes of insect viruses will be studied using single crystal x-ray diffraction and other biophysical methods. The structure of black beetle virus (BBV) will be refined and as many as 3 other related viruses (all in the nodavirus family) will be investigated. The x-ray investigation of these viruses is part of a multigroup effort to relate genomic and protein primary structure (Paul Kaesberg's group, University of Wisconsin, Madison) and high resolution tertiary structure (this proposal) to antigenic and functional behavior of nodavirus serotypes (Roland Rueckert's group, University of Wisconsin, Madison). Each virus displays a distinct phenotype which for at least 3 of the 4 is dependent only on the capsid protein. Tertiary structures will be determined in an attempt to identify functionally important changes in the primary structures studied by Kaesberg. Initially, hypotheses relating structure to stability can be tested using other physical methods. Hypotheses concerned with biological function such as receptor binding, plaque size and cytolytic behavior can be tested by Kaesberg and collaborators who can genetically engineer site specific mutations in the serotype III (FHV) genome, which has been cloned. The RNA transcript from this clone has been shown to be infectious when used as an inoculum with Drosophila Line 1 cells. The long term multidiscipline goal of this project is to produce engineered viruses with each of the phenotypes described in Table I using the FHV clone as the starting gene. The second goal of this proposal and of equal priority to the first is to solve the structure of the insect pathogen Nudaurelia capensis B virus (NBV), the type member of the only class of viruses displaying T-4 quasiequivalent icosahedral symmetry. The primary objective is to identify the mechanism for mediating quasiequivalent interactions and to compare this with T=3 structures (such as black beetle virus) determined at high resolution. The second objective is to compare the subunit structure of NBV with other viral subunits. The 65Kd, tubular monomer is superficially different from the simple or even elaborated eight stranded antiparallel B-barrel structures observed in other viruses.